An effective malaria vaccine will represent a cost-effective and sustainable addition to the currently available malaria control interventions. Malaria vaccines have attempted to target the different stages of malaria infection, typically referred to as the “sporozoite stage”, the “liver stage” and “blood-stage”. The liver stage occurs when sporozoites infect host hepatocytes, multiplying asexually and asymptomatically for a period of 8-30 days. Once in the liver, these organisms differentiate to yield thousands of merozoites which, following rupture of their host cells, escape into the blood and infect red blood cells, thus beginning the erythrocytic or “blood-stage” of the life cycle. Within the red blood cells, the parasites multiply further, again asexually, periodically breaking out of their hosts to invade fresh red blood cells. Several such amplification cycles occur. Thus, classical descriptions of waves of fever arise from simultaneous waves of merozoites escaping and infecting red blood cells.
Anti-asexual blood-stage vaccines have been aimed at reducing parasite growth and multiplication in the blood and, hence, the occurrence or severity of symptoms. Such vaccines should reduce morbidity and mortality due to malaria, in the most susceptible groups (e.g. young children and pregnant women) living in areas where malaria is endemic.
One approach has been to trial recombinant subunit vaccines against blood-stage malaria, but to date none of the subunit blood-stage vaccine candidates have progressed beyond Phase III trials.
An alternative approach has been to use whole blood-stage parasites (e.g. merozoites). Whole blood-stage parasites were first administered to monkeys in 1948 when Freund attempted to develop a whole parasite vaccine. Over the following 30 years there were in excess of 12 separate studies in monkeys (summarized in McCarthy and Good, 2010) until malaria antigens were first cloned in 1983, which ushered in the subunit paradigm era. The whole parasite strategy was abandoned for blood-stage vaccine development because very large numbers of parasites were thought to be required (5×107-1010), making it impossible to prepare such a vaccine at scale in human blood. The only adjuvant shown to be useful was the human-incompatible complete Freund's adjuvant.
More recently, a “low-dose whole parasite” approach has been attempted which may be characterized as follows: (i) it aims to induce a cellular (T cell) immune response, as opposed to an antibody response; (ii) very low doses of parasites are not only sufficient but essential to keep doses low; and (iii) heterologous immunity is induced because presumably the target antigenic determinants of T cells are highly conserved between Plasmodium strains and species (Pinzon-Charry et al, 2010), thereby obviating one of the major impediments of sub-unit vaccines that aim to induce antibodies that target polymorphic merozoite or infected red cell surface proteins. Furthermore, very low doses of parasites overcome the logistic impediment of finding sufficient blood to prepare a vaccine at scale or the need to find a way to grow the parasites in axenic culture. The two critical factors of parasite dose and cellular immune response are in fact closely related. In order to induce a strong cellular immune response it is important to use only a very low dose of parasites (Elliott et al, 2005).
Three studies are reported in which humans or mice have been immunized with ultra low doses of whole parasites—two in which vaccines received an ultra low dose of live infected red cells (with infection terminated by drug early before parasites were visible in the blood by microscopy; Elliott et al, 2005; Pombo et al, 2002), and the other in which mice were immunized with 100-1000 killed infected red cells mixed with CpG and Alum for the primary immunization (Pinzon-Charry et al, 2010). In both humans and mice the immune responses were characterized by a strong in vitro proliferative response to parasites of CD4++/− CD8+ T cells, secretion of γ-interferon and induction of nitric oxide synthase in peripheral blood mononuclear cells. Parasite-specific antibodies were either not induced, or induced at very low levels. Memory T cells (both ‘central’ and ‘effector’) were induced. It has been proposed that induced memory T cells will have specificity for internal antigens of the parasite and that these will be highly conserved as they are not under immune selective pressure from B cells and antibody. This has been shown to be the case for the one major T cell target antigen of Plasmodium falciparum identified so far, the purine salvage enzyme, HGXPRT (Makobongo et al, 2003) where immunity appeared to be mediated solely by T cells.
The requirement for purine nucleotide salvage by blood-stage malaria parasites has been exploited by making genetically-attenuated malaria parasites lacking a functional gene encoding the purine nucleotide transporter 1 (Pynt1−), which upon administration to mice provided sterile immunity against subsequent malaria infection (Ahmed et al., 2010; International Publication 2008/094183). However, the applicability of this approach to humans is questionable because of the possibility of selection of genetic “breakout” following administration to humans.